Compositions and methods for treating metabolic disorders

ABSTRACT

Disclosed are methods for treating a subject suffering from phenylketonuria and/or phenylalanemia. The methods include, in part, enterally administering to the subject a LNAA supplement in which the weight ratio of Leu to Val is greater than 2:1; in which the weight ratio of Leu to iLeu is greater than 3:1; or which includes one or more LNAAs and which further includes Lys. LNAA supplements are also disclosed. Also disclosed are methods for treating a subject suffering from a condition involving a metabolic disorder involving the metabolism of a first amino acid X. The method includes enterally administering to the subject a composition which (i) is substantially free from the first amino acid X and (ii) which includes a second amino acid Y that competes with amino acid X at a gastrointestinal tract transporter.

CROSS REFERENCE TO RELATED APPLICATIONS

This application is a Continuation application of U.S. patentapplication Ser. No. 12/805,554, filed Aug. 5, 2010, which is aDivisional application of U.S. patent application Ser. No. 10/826,112,filed Apr. 17, 2004, now abandoned, which claims the benefit of U.S.Provisional Patent Application Ser. No. 60/490,473, filed Jul. 28, 2003,all of which are hereby incorporated in their entireties by reference.

FIELD OF THE INVENTION

The subject invention is directed, generally, to methods and materialsfor treating conditions associated with metabolic disorders and, moreparticularly, to methods and materials for treating conditionsassociated with metabolic disorders of particular amino acids.

BACKGROUND OF THE INVENTION

A number of conditions which afflict humans and other animals areattributable to disorders in metabolizing particular amino acids. Inmany of these conditions, treatment involves .restricting the dietaryintake of the particular amino acid or amino acids associated with thecondition. However, therapies based on dietary restriction requirepatient compliance and also requires that the patient know whether aparticular food contains the particular amino acid or amino acidsassociated with the condition.

For example, phenylketonuria (“PKU”) is hyperaminoacidemia ofphenylalanine (Phe) associated with an inborn error of phenylalaninemetabolism, mutation of the gene encoding phenylalanine 4-hydroxylase(“PAH”), which converts phenylalanine to tyrosine. In some cases, anadditional metabolic defect occurs in the synthetic pathway of eitherdihydropteridine or tetrahydrobiopterin (“BH4”), PAH co-factors,contributing further to the hyperphenylalaninemia (“HPA”). Whereas anormal plasma Phe level is approximately 0.05 mM (Pardridge, “BloodBrainBarrier Amino Acid Transport: Clinical Implications,” chapter 6 inInborn Errors of Metabolism in Humans, Cockburn et al., eds, Lancaster,England: MTP Press Ltd. (1980) (“Pardridge”), untreated “classic” PKUpatients have plasma Phe levels above 1 mM (e.g., plasma Phe levels offrom about 1 mM to about 2.5 mM or more), and, although treatment with alow-Phe diet has a goal of reducing plasma Phe to below 0.3 mM, this isdifficult to attain due to dietary compliance problems. In the US, 1 in10,000 babies are born with PKU.

The excessive levels of plasma phenylalanine observed in PKU combinedwith the relatively high affinity of Phe for binding sites on carrierprotein of the neutral amino acid transport system in the bloodbrainbarrier (“BBB”) leads to (i) accumulation of Phe and its neurotoxicmetabolites (e.g., phenylpyruvate, phenylacetate, phenyllactate) in thebrain and (ii) depressed levels of non-Phe neutral amino acids enteringthe brain, resulting in disturbed brain development and function, sincekey cerebral pathways of metabolism (e.g., synthesis ofneurotransmitters) require precursor amino acids, such as tyrosine. Thisdepression is pronounced for tyrosine, which is low in the plasma supplydue to the PKU metabolic error in the enzyme responsible for convertingphenylalanine to tyrosine. Current thought is that the neurologicaldeficits of PKU are due predominantly to the depression of levels ofnon-Phe neutral amino acids entering the brain (Kaufman, “Some FactsRelevant to a Consideration of a Possible Alternative Treatment forClassical Phenylketonuria,” J. Inher. Metab. Dis., 21 (supplement 3):4-19 (1998) (“Kaufman”)).

Although a diet low in phenylalanine can reduce plasma Phe levels in“classic” PKU below 0.3 mM and ameliorate the mental retardationassociated with untreated PKU, dietary compliance becomes problematic asPKU patients reach adolescence, leading to a rise in plasma Phe levelsand to both loss in intelligence and white matter changes in the brain.Nutritional deficiencies can also result from Phe restricted diets.Alternative treatments have thus been developed. For example, toovercome suspected depletion of the neurotransmitters dopamine andserotonin, PKU patients have been treated with the neurotransmitterprecursors tyrosine and tryptophan (Lou, “Large Doses of Tryptophan andTyrosine as Potential Therapeutic Alternative to Dietary PhenylalanineRestriction in Phenylketonuria,” Lancet, 2:150-151 (1983)). To reduceinflux of Phe into the brain, a supplement of branched chain neutralamino acids containing valine, isoleucine, and leucine, was administeredto older PKU patients (Berry et al, “Valine, Isoleucine and Leucine. ANew Treatment for Phenylketonuria,” Am. J. Dis. Child., 144:539-543(1990) (“Berry”), who reported significant improvement in behavioraldeficits. In Kaufman, it was proposed that the addition of theneurotransmitter precursors, tyrosine and tryptophan to Berry'ssupplement, should lead to further improvement. However, efficacy ofthese dietary amino acid supplement treatments has been controversial.

Tyrosinemia is another example of a condition that is attributable to adisorder in metabolizing particular amino acids. More particularly,tyrosinemia is a disorder caused by a defect in the terminal enzyme ofthe tyrosine metabolic pathway, leading to accumulation offumarylacetoacetate, which converts to succinylacetone, whichaccumulates and is toxic to the liver. Tyrosinemia is associated withliver failure, liver diseases, and hepatocarcinoma. Livertransplantation can restore normal enzyme activity to the tyrosinemetabolic pathway and is utilized in advanced cases. However this is adifficult and expensive therapy. Another currently employed therapy fortyrosinemia includes a two-fold approach: (i) use of a new inhibitor oftyrosine hydroxylase,.NTBC((2-(2-nitro-4-trifluoromethylbenzoyl)-1,3-cyclohexanedione), whichprevents formation of succinylacetone; and (ii) a diet low in bothtyrosine and phenylalanine to manage the amount of tyrosine which mustbe metabolized. However, safety issues regarding NTBC are unanswered todate, and dietary restriction of tyrosine and phenylalanine is dependenton patient knowledge and compliance, which, as mentioned above, can beproblematic, especially in adolescents and adults.

Alkaptonuria is another example of a condition that is attributable to adisorder in metabolizing particular amino acids. Current therapiesinclude restricting dietary intake of phenylalanine and tyrosine toreduce accumulation of the metabolite, homogentisic acid. Some patientstake NTBC and vitamin C to reduce homogentisic acid aggregates. However,safety issues regarding NTBC are unanswered to date, and dietaryrestriction of tyrosine and phenylalanine is dependent on patientknowledge and compliance, which, as mentioned above, can be problematic.

Homocystinuria is another example of a condition that is attributable toa disorder in metabolizing particular amino acids. Patients with thiscondition frequently follow a methionine restricted diet. However,dietary restriction of methionine is dependent on patient knowledge andcompliance, which, as mentioned above, can be problematic.

A number of conditions are attributable to metabolic disorders affectingthe metabolism of the branched chain amino acids (“BCAAs”), such asleucine, isoleucine, and valine. Leucine, isoleucine, and valine areessential amino acids which must be obtained from dietary protein. Adefect in one step of a multistep metabolic pathway which converts theBCAAs to energy, results in accumulation of an intermediate metaboliteof the BCAA to toxic levels, causing disease. This is a large group ofdiseases that includes, for example, maple syrup urine disease (“MSUD”),isovaleric acidemia, methylmalonic acidemia, and propionic acidemia.These diseases are treated with special dietary formulas low in the BCAAhaving the metabolic defect. However, as discussed above, successfulmanagement of such diseases and conditions by dietary restriction of aparticular amino acid or a particular set of amino acids is dependent onpatient knowledge and compliance, which can be problematic.

In view of the above, there is a need for methods and materials fortreating conditions, such as phenylketonuria, that are attributable to adisorder in metabolizing particular amino acids, and the presentinvention, in part, is directed to meeting this need.

SUMMARY OF THE INVENTION

The present invention relates to a method for treating a subjectsuffering from phenylketonuria and/or phenylalanemia. This methodincludes enterally administering to the subject a LNAA supplement inwhich the weight ratio of Leu to Val is greater than 2:1.

The present invention also relates to another method for treating asubject suffering from phenylketonuria and/or phenylalanemia. Thismethod includes enterally administering to the subject a LNAA supplementin which the weight ratio of Leu to iLeu is greater than 3:1.

The present invention also relates to another method for treating asubject suffering from phenylketonuria and/or phenylalanemia. Thismethod includes enterally administering to the subject a LNAA supplementwhich comprises one or more .LNAAs and which further comprises Lys.

The present invention also relates to a LNAA supplement which includesLeu and Val and in which the weight ratio of Leu to Val is greater than2:1.

The present invention also relates to a LNAA supplement which includesLeu and iLeu and in which the weight ratio of Leu to iLeu is greaterthan 3:1.

The present invention also relates to a LNAA supplement which includesone or more LNAAs and further includes Lys.

The present invention also relates to a LNAA supplement which includes,per 600 mg of LNAA supplement, from about 100 mg to about 290 mg of Tyr;from about 30 mg to about 90 mg of Trp; from about 25 mg to about 75 mgof Met; from about 15 mg to about 45 mg of iLeu; from about 15 mg toabout 50 mg of Threo; from about 15 mg to about 50 mg of Val; from about40 mg to about 200 mg of Leu; from about 15 mg to about 45 mg of His;and from about 15 mg to about 50 mg of Arg.

The present invention also relates to a method for treating a subjectsuffering from a condition involving a metabolic disorder involving themetabolism of a first amino acid X. The method includes enterallyadministering to the subject a composition which (i) is substantiallyfree from the first amino acid X and (ii) which includes a second aminoacid Y that competes with amino acid X at a gastrointestinal tracttransporter. As one skilled in the art will appreciate, the compositioncan further include (i.e., in addition to second amino acid Y) othercomponents, such as other amino acids (e.g., one or more other aminoacids which compete with amino acid X at a gastrointestinal tracttransporter).

DETAILED DESCRIPTION OF THE INVENTION

As used herein, “LNAA supplement” is meant to refer to any compositionwhich includes, at a minimum, one or more large neutral amino acids,such as Phe, Leu, Tyr, Trp, Met, iLeu, Val, and Threo. The LNAAsupplement can optionally include other components, such as basic aminoacids (e.g., Arg, His, Lys, etc.) and/or other amino acids, vitamins,minerals, binders, diluents, dispersing agents, and other excipients.Illustratively, the LNAA supplement can include one, two, three, four,five, six, or more than six large neutral amino acids. The LNAAsupplement can be substantially free from one or more specified aminoacids, as in the case, for example, where the LNAA supplement issubstantially free from amino acid Z. As used in this context, an LNAAsupplement is to be deemed to be substantially free from amino acid Zwhen amino acid Z is present in an amount that is less than 10% (e.g.,less than about 9%, less than about 8%, less than about 7%, less thanabout 6%, less than about 5%, less than about 4%, less than about 3%,less than about 2%, and/or less than about 1%), by weight, of the totalweight of all of the large neutral amino acids present in the LNAAsupplement.

“Treating”, as used herein, is meant to refer to treatment of the director indirect cause of a condition; to treatment of a condition'ssymptoms; or to both.

“Subject”, as used herein, is meant to refer to any animal, such as anymammal, e.g., mice rats, cats, rabbits, dogs, pigs, horses, cows, andprimates, such as humans. Illustratively, “subject”, as used herein, ismeant to include human infants, human children, human adolescents, humanadults, male humans, female humans, humans who are less than about 2years of age, humans who are between about 2 years of age and 5 years ofage, humans who are between about 5 and about 10 years of age, humanswho are between about 10 and about 18 years of age, humans who arebetween about 18 and about 30 years of age, humans who are between about30 and about 40 years of age, humans who are between about 40 and about50 years of age, humans who are between about 50 and about 60 years ofage, humans who are over about 60 years of age, humans suffering fromphenylketonuria, humans not suffering from phenylketonuria, humanssuffering from phenylalanemia, humans not suffering from phenylalanemia,humans suffering from tyrosinemia, humans not suffering fromtyrosinemia, humans suffering from alkaptonuria, humans not sufferingfrom alkaptonuria, humans suffering from homocystinuria, humans notsuffering from homocystinuria, humans suffering from maple syrup urinedisease, humans not suffering from maple syrup urine disease, humanssuffering from isovaleric acidemia, humans not suffering from isovalericacidemia, humans suffering from methylmalonic acidemia, humans notsuffering from methylmalonic acidemia, humans suffering from propionicacidemia, and/or humans not suffering from methylmalonic acidemia.

As used herein, “enteral administration” of a substance is meant torefer to any administration which delivers the substance to one or moreportions of the gastrointestinal (“GI”) tract, such as the stomach, thesmall intestine, and the large intestine. For example, enteraladministration can be carried out orally, for example, by swallowing atablet, capsule, or other solid dosage form or by swallowing a liquidsolution or suspension. Additionally or alternatively, enteraladministration can be carried out by feeding tube, by gavage, or byother common methods of enteral administration.

One aspect of the present invention relates to a method for treating asubject suffering from phenylketonuria and/or phenylalanemia. Thismethod includes enterally administering to the subject a LNAA supplementin which the weight ratio of Leu to Val is greater than 2:1, such asgreater than about 2.2:1, greater than about 2.5:1, greater than about2.8:1, greater than about 3:1, greater than about 3.2:1, greater thanabout 3.5:1, greater than about 3.8:1, greater than about 4:1, greaterthan about 4.2:1, greater than about 4.5:1, greater than about 4.8:1,greater than about 5:1, greater than about 5.2:1, greater than about5.5:1, greater than about 5.8:1, greater than about 6:1, and/or greaterthan about 6.2:1. The LNAA supplement can be substantially free fromphenylalanine. The LNAA supplement can also include Arg but no His orLys; it can also include His but no Arg or Lys; it can also include Lysbut no His or Arg; it can also include Arg and Lys but no His; it canalso include Arg and His but no Lys; it can also include His and Lys butno Arg; or it can also include His, Arg, and Lys.

Another aspect of the present invention relates to a method for treatinga subject suffering from phenylketonuria and/or phenylalanemia. Thismethod includes enterally administering to the subject a LNAA supplementin which the weight ratio of Leu to iLeu is greater than 3:1, such asgreater than about 3.2:1, greater than about 3.5:1, greater than about3.8:1, greater than about 4:1, greater than about 4.2:1, greater thanabout 4.5:1, greater than about 4.8:1, greater than about 5:1, greaterthan about 5.2:1, greater than about 5.5:1, greater than about 5.8:1,greater than about 6:1, and/or greater than about 6.2:1. This LNAAsupplement can be one in which the weight ratio of Leu to Val in theLNAA supplement is greater than 2:1, such as in the case where theweight ratio of Leu to Val is greater than about 2.2:1, greater thanabout 2.5:1, greater than about 2.8:1, greater than about 3:1, greaterthan about 3.2:1, greater than about 3.5:1, greater than about 3.8:1,greater than about 4:1, greater than about 4.2:1, greater than about4.5:1, greater than about 4.8:1, greater than about 5:1, greater thanabout 5.2:1, greater than about 5.5:1, greater than about 5.8:1, greaterthan about 6:1, and/or greater than about 6.2:1. The LNAA supplement canbe substantially free from phenylalanine. The LNAA supplement can alsoinclude Arg but no His or Lys; it can also include His but no Arg orLys; it can also include Lys but no His or Argi it can also include Argand Lys but no His; it can also include Arg and His but no Lys; it canalso include His and Lys but no Arg; or it can also include His, Arg,and Lys.

Another aspect of the present invention relates to a method for treatinga subject suffering from phenylketonuria and/or phenylalanemia. Thismethod includes enterally administering to the subject a LNAA supplementwhich includes one or more LNAAs and which further includes Lys.

Illustratively, the LNAA supplement of this aspect of the presentinvention can include Leu and Lys; or the LNAA supplement can includeLeu and iLeu and Lys, such as where the LNAA supplement includes Leu andiLeu and Lys and where the weight ratio of Leu to iLeu in the LNAAsupplement is greater than about 0.5:1, greater than about 1:1, greaterthan 1:1, greater than about 1.5:1, greater than about 2:1, greater thanabout 2:1, greater than about 2.5:1, greater than about 3:1, greaterthan 3:1, greater than about 3.2:1, greater than about 3.5:1, greaterthan about 3.8:1, greater than about 4:1, greater than about 4.2:1,greater than about 4.5:1, greater than about 4.8:1, greater than about5:1, greater than about 5.2:1, greater than about 5.5:1, greater thanabout 5.8:1, greater than about 6:1, and/or greater than about 6.2:1.

As further illustration, the LNAA supplement of this aspect of thepresent invention can include Leu and Lys; or the LNAA supplement caninclude Leu and Val and Lys, such as where the LNAA supplement includesLeu and Val and Lys and where the weight ratio of Leu to Val in the LNAAsupplement is greater than about 0.5:1, greater than about 1:1, greaterthan 1:1, greater than about 1.5:1, greater than about 2:1, greater thanabout 2:1, greater than about 2.5:1, greater than about 3:1, greaterthan 3:1, greater than about 3.2:1, greater than about 3.5:1, greaterthan about 3.8:1, greater than about 4:1, greater than about 4.2:1,greater than about 4.5:1, greater than about 4.8:1, greater than about5:1, greater than about 5.2:1, greater than about 5.5:1, greater thanabout 5.8:1, greater than about 6:1, and/or greater than about 6.2:1.

As yet further illustration, the LNAA supplement of this aspect of thepresent invention can be one which includes Leu; in which the weightratio of Leu to iLeu in the LNAA supplement is greater than about 0.5:1;and in which the weight ratio of Leu to Val in the LNAA supplement isgreater than about 0.5:1. For example, the LNAA supplement of thisaspect of the present invention can be one which includes Leu; in whichthe weight ratio of Leu to iLeu in the LNAA supplement is greater than3:1; and in which the weight ratio of Leu to Val in the LNAA supplementis greater than about 0.5:1. As another example, the LNAA supplement ofthis aspect of the present invention can be one which includes Leu; inwhich the weight ratio of Leu to iLeu in the LNAA supplement is greaterthan about 0.5:1; and in which the weight ratio of Leu to Val in theLNAA supplement is greater than 2:1. As yet another example, the LNAAsupplement of this aspect of the present invention can be one whichincludes Leu; in which the weight ratio of Leu to iLeu in the LNAAsupplement is greater than 3:1; and in which the weight ratio of Leu toVal in the LNAA supplement is greater than 2:1.

As one skilled in the art will appreciate, the methods of this aspect ofthe present invention (i.e., by enterally administering to the subject aLNAA supplement which includes one or more LNAAs and which furtherincludes Lys) can be practiced with an LNAA supplement that issubstantially free from phenylalanine. Additionally or alternatively,the LNAA supplement can further include Arg and/or His or the LNAAsupplement can be substantially free from Arg and/or His.

In one particular embodiment of this aspect of the present invention,the LNAA supplement includes, per 500 mg of LNAA supplement:

from about 100 mg to about 290 mg of Tyr;

from about 25 mg to about 75 mg of Trp;

from about 15 mg to about 50 mg of Met;

from about 15 mg to about 55 mg of iLeu;

from about 15 mg to about 50 mg of Threo;

from about 15 mg to about 55 mg of Val;

from about 15 mg to about 200 mg of Leu;

from about 10 mg to about 30 mg of His; and

from about 5 mg to about 200 mg of Lys.

As one skilled in the art will appreciate, the amounts of each aminoacid present in this LNAA supplement can be varied within the statedlimits. Thus, for example, the LNAA supplement can include, per 500 mgof LNAA supplement, from about 10 mg to about 30 mg of Lys. Moreover, asone skilled in the art will also appreciate, this LNAA supplement canoptionally include one or more other amino acids not mentioned above, orthe LNAA supplement can be substantially free of one or more other aminoacids not mentioned above. Thus, for example, this LNAA supplement canoptionally include Arg, or it can be substantially free from arginine;and/or it can be substantially free from phenylalanine

Yet another aspect of the present invention relates to a method fortreating a subject suffering from phenylketonuria and/or phenylalanemia.This method includes enterally administering to the subject a LNAAsupplement which includes, per 600 mg of LNAA supplement:

from about 100 mg to about 290 mg of Tyr;

from about 30 mg to about 90 mg of Trp;

from about 25 mg to about 75 mg of Met;

from about 15 mg to about 45 mg of iLeu;

from about 15 mg to about 50 mg of Threo;

from about 15 mg to about 50 mg of Val;

from about 40 mg to about 200 mg of Leu;

from about 15 mg to about 45 mg of His; and

from about 15 mg to about 50 mg of Arg.

As one skilled in the art will appreciate, the amounts of each aminoacid present in this LNAA supplement can be varied within the statedlimits. Moreover, as one skilled in the art will also appreciate, theLNAA supplement of this aspect of the present invention can optionallyinclude one or more other amino acids not mentioned above, or the LNAAsupplement of this aspect of the present invention can be substantiallyfree of one or more other amino acids not mentioned above. Thus, forexample, the LNAA supplement of this aspect of the present invention canoptionally include Lys (e.g., from about 5 mg to about 200 mg of Lys per600 mg of LNAA supplement, or it can be substantially free from Lys;and/or it can be substantially free from phenylalanine

The methods of the various aspects of the present invention discussedabove can be carried out by any suitable form of enteral administrationof the LNAA supplement to the subject. It will be appreciated that theactual preferred amount of LNAA supplement to be administered accordingto the present invention will vary according to the particular largeneutral amino acid or acids that are present in the LNAA supplement, thenature of the other components present in the LNAA supplement, and theform of enteral of administration. Many factors that may modify theaction of the LNAA supplement (e.g., body weight, sex, diet, time ofadministration, route of administration, rate of excretion, condition ofthe subject, drug combinations, and reaction sensitivities andseverities) can be taken into account by those skilled in the art.Administration can be carried out continuously or periodically withinthe maximum tolerated dose. Optimal administration rates for a given setof conditions can be ascertained by those skilled in the art usingconventional dosage administration tests, such as those described in theexamples which follow. Briefly, dosing can be based on the level ofplasma phenylalanine For example, dosing can be based on the level ofplasma phenylalanine at 0, 3, and/or 6 hours following administration ofthe LNAA supplement.

Illustratively, the LNAA supplement can be administered in a single oraldose of from about 0.1 g to about 10 g per kg of the subject's bodyweight substantially at mealtime. As used herein, “substantially atmealtime” is meant to refer to the period of time from about 4 hoursbefore mealtime to about 1 hour after mealtime, such as from about 4hours before mealtime to about mealtime, from about 3 hours beforemealtime to about mealtime, from about 2 hours before mealtime to aboutmealtime, from about 1 hour before mealtime to about mealtime, and/orfrom about 0.5 hours before mealtime to about mealtime. For example, theLNAA supplement can be administered in a single oral dose of from about0.2 g to about 5 g per kg, such as from about 0.3 g to about 3 g per kg,from about 0.4 g to about 2 g per kg, from about 0.5 g to about 1 g perkg of the subject's body weight substantially at mealtime.Alternatively, the LNAA supplement can be administered, for example, inmultiple oral doses spaced throughout the day (e.g., administered every2-6 hours). Optionally, the LNAA supplement can be formulated so as toprovide sustained release of the LNAAs over a period of time.

The methods of the various aspects of the present invention discussedabove can include further steps. Illustratively, the method of thepresent invention can further include restricting the subject's dietaryintake of phenylalanine. For the purposes of the present invention, asubject's dietary intake of phenylalanine is to be deemed to berestricted if the subject's diet (i) is chosen, in whole or in part, onthe basis of phenylalanine content or (ii) if the subject's dietcontains a total daily phenylalanine intake substantially less (e.g.,more than 50% less) than the general population's total dailyphenylalanine intake. Alternatively, the method of the present inventioncan further include not restricting the subject's dietary intake ofphenylalanine. For the purposes of the present invention, a subject'sdietary intake of phenylalanine is to be deemed to be not restricted ifthe subject's diet contains a total daily phenylalanine intake that issubstantially the same (e.g., plus or minus less than 50%) as thegeneral population's total daily phenylalanine intake.

The present invention, in yet other aspects thereof, relates to theaforementioned LNAA supplements.

Illustratively, the present invention relates to a LNAA supplement whichincludes Leu and Val and in which the weight ratio of Leu to Val isgreater than 2:1.

In another aspect, the present invention relates to a LNAA supplementwhich includes Leu and iLeu and in which the weight ratio of Leu to iLeuis greater than 3:1.

In yet another aspect, the present invention relates to a LNAAsupplement which includes one or more LNAAs and further includes Lys.

In still another aspect, the present invention relates to a LNAAsupplement which includes, per 600 mg of LNAA supplement, from about 100mg to about 290 mg of Tyr; from about 30 mg to about 90 mg of Trp; fromabout 25 mg to about 75 mg of Met; from about 15 mg to about 45 mg ofiLeu; from about 15 mg to about 50 mg of Threo; from about 15 mg toabout 50 mg of Val; from about 40 mg to about 200 mg of Leu; from about15 mg to about 45 mg of His; and from about 15 mg to about 50 mg of Arg.

Examples of the LNAA supplements of the present invention include thosedescribed hereinabove with reference to the methods of the presentinvention. These LNAA supplements can be prepared using amino acidsderived from natural sources, or the amino acids can be preparedsynthetically by methods well known to those skilled in the art. Theycan be of any suitable dosage form, such as those discussed above,suitable for enteral administration, and they can contain, in additionto large neutral amino acids and other amino acids, vitamins, minerals,excipients, and the like. For example, suitable dosage forms for oraladministration include tablets, dispersible powders, granules, capsules,suspensions, syrups, and elixirs. Inert diluents and carriers fortablets include, for example, calcium carbonate, sodium carbonate,lactose, and talc. Tablets may also contain granulating anddisintegrating agents, such as starch and alginic acid; binding agents,such as starch, gelatin, and acacia; and lubricating agents, such asmagnesium stearate, stearic acid, and talc. Tablets may be uncoated ormay be coated by known techniques to delay disintegration andabsorption. Inert diluents and carriers which may be used in capsulesinclude, for example, calcium carbonate, calcium phosphate, and kaolin.The aforementioned capsules or tablets can, optionally, be formulated toas to provide sustained release of the LNAAs over a period of time.Suspensions, syrups, and elixirs may contain conventional excipients,for example, methyl cellulose, tragacanth, sodium alginate; wettingagents, such as lecithin and polyoxyethylene stearate; andpreservatives, such as ethyl-p-hydroxybenzoate.

The present invention, in yet another aspect thereof, relates to amethod for treating a subject suffering from a condition involving ametabolic disorder involving the metabolism of a first amino acid X. Themethod includes enterally administering to the subject a compositionwhich is substantially free from said first amino acid X and whichincludes a second amino acid Y that competes with amino acid X at agastrointestinal tract transporter, such as a Caco-2 cell transporter.As used in this context, a composition is to be deemed to besubstantially free from first amino acid X when amino acid X is presentin an amount that is less than 10% (e.g., less than about 9%, less thanabout 8%, less than about 7%, less than about 6%, less than about 5%,less than about 4%, less than about 3%, less than about 2%, and/or lessthan about 1%), by weight, of the total weight of all of the amino acidspresent in the composition.

As one skilled in the art will appreciate, the composition used in thisaspect of the present invention can further include (i.e., in additionto second amino acid Y) other components, such as other amino acids(e.g., one or more other amino acids which compete with amino acid X ata Caco-2 cell transporter or other gastrointestinal tract transporter).Moreover, as one skilled in the art will appreciate, the method of thisaspect of the present invention can further include additional steps,such as administering a second composition which includes, for example,one or more other amino acids (e.g., other than second amino acid Y)which compete with amino acid X at a Caco-2 cell transporter or anothergastrointestinal tract transporter.

In one embodiment of this aspect of the present invention, the conditionis phenylketonuria and/or phenylalanemia, the first amino acid X isphenylalanine, and the composition is one of the LNAA supplementsdescribed hereinabove.

In another embodiment of this aspect of the present invention, thecondition is not phenylketonuria and/or phenylalanemia.

In another embodiment of this aspect of the present invention, thecondition is tyrosinemia; the first amino acid X is tyrosine; and thesecond amino acid Y is selected from Phe, Leu, Trp, Lys, His, andcombinations thereof.

In yet another embodiment of this aspect of the present invention, thecondition is tyrosinemia; the first amino acid X is tyrosine; and thesecond amino acid Y is selected from Leu, Trp, Lys, His, andcombinations thereof.

In still another embodiment of this aspect of the present invention, thecondition is alkaptonuria; the first amino acid X is selected fromphenylalanine, tyrosine, and combinations thereof; and the second aminoacid Y is selected from Leu, Trp, Lys, His, and combinations thereof.

In still another embodiment of this aspect of the present invention, thecondition is homocystinuria; wherein the first amino acid X ismethionine; and the second amino acid Y is an amino acid that competeswith methionine at a gastrointestinal tract transporter.

In still other embodiments of this aspect of the present invention, thecondition is a disorder affecting metabolism of a branched amino acidselected from leucine, isoleucine, valine, and combinations thereof; thefirst amino acid X is selected from leucine, isoleucine, valine, andcombinations thereof; and the second amino acid Y is an amino acid thatcompetes with the first amino acid X at a gastrointestinal tracttransporter. Such conditions include, for example, maple syrup urinedisease, isovaleric acidemia, methylmalonic acidemia, and propionicacidemia.

The method of this aspect of the present invention can be carried out byany suitable form of enteral administration of the composition to thesubject. It will be appreciated that the actual preferred amount ofcomposition to be administered according to the present invention willvary according to the particular amino acid or acids that are present inthe composition, the nature of the other components present in thecomposition, and the form of enteral of administration. Many factorsthat may modify the action of the composition (e.g., body weight, sex,diet, time of administration, route of administration, rate ofexcretion, condition of the subject, drug combinations, and reactionsensitivities and severities) can be taken into account by those skilledin the art. Administration can be carried out continuously orperiodically within the maximum tolerated dose. Optimal administrationrates for a given set of conditions can be ascertained by those skilledin the art using conventional dosage administration tests, such as thosedescribed in the examples which follow. Briefly, dosing can be based onthe plasma level of first amino acid X. For example, dosing can be basedon the plasma level of first amino acid X at 0, 3, and/or 6 hoursfollowing administration of the composition.

Illustratively, the composition can be administered in a single oraldose of from about 0.1 g to about 10 g per kg of the subject's bodyweight substantially at mealtime. For example, the composition can beadministered in a single oral dose of from about 0.2 g to about 5 g perkg, such as from about 0.3 g to about 3 g per kg, from about 0.4 g toabout 2 g per kg, from about 0.5 g to about 1 g per kg of the subject'sbody weight substantially at mealtime. Alternatively, the compositioncan be administered, for example, in multiple oral doses spacedthroughout the day (e.g., administered every 2-6 hours).

The method this aspect of the present invention can include furthersteps (i.e., in addition to enteral administration of the composition.Illustratively, the method of the present invention can further includerestricting the subject's dietary intake of first amino acid X. For thepurposes of the present invention, a subject's dietary intake of firstamino acid X is to be deemed to be restricted if the subject's diet (i)is chosen, in whole or in part, on the basis of the amount of firstamino acid X present in a particular food or (ii) if the subject's dietcontains a total daily intake of first amino acid X that issubstantially less (e.g., more than 50% less) than the generalpopulation's total daily intake of first amino acid X. Alternatively,the method of the present invention can further include not restrictingthe subject's dietary intake of first amino acid X. For the purposes ofthe present invention, a subject's dietary intake of first amino acid Xis to be deemed to be not restricted if the subject's diet contains atotal daily intake of first amino acid X that is substantially the same(e.g., plus or minus less than 50%) as the general population's totaldaily intake of first amino acid X.

Compositions useful in the practice of this aspect of the presentinvention can be prepared using amino acids derived from naturalsources, or the amino acids can be prepared synthetically by methodswell know to those skilled in the art. The compositions can be of anysuitable dosage form, such as those discussed above, suitable forenteral administration, and they can contain, in addition to amino acidY, other amino acids, vitamins, minerals, excipients, and the like. Forexample, suitable dosage forms for oral administration include tablets,dispersible powders, granules, capsules, suspensions, syrups, andelixirs. Inert diluents and carriers for tablets include, for example,calcium carbonate, sodium carbonate, lactose, and talc. Tablets may alsocontain granulating and disintegrating agents, such as starch andalginic acid; binding agents, such as starch, gelatin, and acacia; andlubricating agents, such as magnesium stearate, stearic acid, and talc.Tablets may be uncoated or may be coated by known techniques to delaydisintegration and absorption. Inert diluents and carriers which may beused in capsules include, for example, calcium carbonate, calciumphosphate, and kaolin. Suspensions, syrups, and elixirs may containconventional excipients, for example, methyl cellulose, tragacanth,sodium alginate; wetting agents, such as lecithin and polyoxyethylenestearate; and preservatives, such as ethyl-p-hydroxybenzoate.

The present invention is further illustrated with the followingexamples. In some of the following examples, applicant discusses what ispresently believed to be the mechanism by which the present inventionoperates. This discussion is presented only for the purposes ofdiscussion and is not meant, in any way, to limit the scope of thepresent invention.

EXAMPLES Example 1 The Hypothesis

The availability of amino acids in the brain is determined by (i) theplasma supply of the amino acid, and (ii) competition of theplasma-supplied amino acids for a common amino acid binding site(s) onthe carrier protein of the BBB neutral amino acid transporter. It hasbeen hypothesized that competition for neutral amino acids at a commoncarrier binding site, under physiological conditions, is unique to thecentral nervous system (Pardridge, which is hereby incorporated byreference), and that such competition is the basis for the correlationof BBB transport and clinical disorders affecting the brain (e.g., PKU).Whereas prior PKU-related studies have focused on competitive transportof non-Phe LNAAs across the blood brain barrier so as to suppress entryof Phe into the brain, it has ignored the transport of LNAAs out of thegastrointestinal tract and into the blood, which can be a majordeterminant of the plasma amino acid supply.

Nine separate transport systems have been identified in the BBB(Oldendorf, “Measurement of Brain Uptake of Radiolabelled SubstancesUsing a Tritiated Water Internal Standard,” Brain Res., 24(2):372-376(1970), which is hereby incorporated by reference). Transport of a givensubstrate across the BBB is characterized by its affinity constant Km. Alower Km value corresponds to greater affinity for the binding site ofthe carrier protein. Each BBB transport system mediates thetrans-capillary flux of a group of substrates. For example, onetransport system mediates transport of LNAAs, another mediates transferof hexoses, etc.

Three of the BBB transport systems mediate transport of the common aminoacids, with separate carrier proteins for LNAAs, for basic amino acids,and for acidic amino acids. The values of the Michaelis constant, Km,for the three classes of common amino acids are presented below in Table1 (Pardridge, which is hereby incorporated by reference).

TABLE 1 representative amino acid or system other substrate Km (mM)neutral amino acids Phe 0.12 basic amino acids Lys 0.10 acidic aminoacids Glu 0.04 hexoses glucose 9 thyroid hormone T3 0.00 1 1

Although much quantitative information has been obtained on the BBBtransport systems, relatively little is known regarding the modulationof the carrier proteins. Developmental or pathological induction orrepression of transporter activity would be expected to profoundlyinfluence the pathways of brain metabolism, which are limited byprecursor availability.

The absolute and apparent Km values of the neutral amino acids at theBBB have been determined experimentally (Pardridge, which is herebyincorporated by reference). The absolute value of Km is the value of Kmin the absence of competition from other neutral amino acids for thebinding site on the LNAA carrier protein. The “apparent Km” is the valueof Km in the presence of other LNAAs competing for the binding site onthe LNAA carrier protein. The apparent Km (“Km(app)”) value of a givenamino acid is calculated from the absolute Km value and the sum of theratios of the plasma level of each LNAA divided by its Km value, asshown in Equation 1, below.

Km(app)=Km(1+Σ[aa]/Km)  (Eq. 1)

The experimental values of Km(app) for the LNAA transport system in theBBB are presented below in Table 2 (Pardridge, which is herebyincorporated by reference).

TABLE 2 typical plasma Km Km(app) amino acid level (mM) (mM) mM) LNAA'sPhe 0.05 0.12 0.45 Leu 0.10 0.15 0.53 Tyr 0.09 0.16 0.58 Trp 0.10 0.190.71 Met 0.04 0.19 0.77 iLeu 0.07 0.33 1.3 Val 0.14 0.63 2.5 Threo 0.190.73 3.0 Basic aa's His 0.05 0.28 1.1 Arg 0.10 0.09 0.40 Lys 0.30 0.100.25Equation 1 predicts that, if the plasma level of an LNAA is much lessthan its value of Km, then that amino acid will not compete effectivelyfor the carrier protein binding site. The Km for binding of LNAAs tocarrier proteins in organs other than brain is 5-10 mM (see Table 3),which is 50-100 times higher than the physiological plasma concentrationof LNAAs. Equation 1 predicts that significant competition effects willnot occur under normal physiological conditions in vivo for LNAAs intissues other than brain. However, competition has been demonstrated inperipheral tissue in vitro at plasma amino acid concentrations of 5-50M. From these observations, applicant hypothesized that high levels ofnon-Phe amino acid supplement could conceivably compete with Phe at theGI tract transporter.

Experimental values of Km for transport in intestinal epithelia arepresented below, in Table 3 (Pardridge, which is hereby incorporated byreference).

TABLE 3 amino acid intestinal epithelia Km (mM) Phe 1 Leu 2 Met 5 His 6Val 3Since LNAAs are associated with several clinical disorders, and sincethe LNAAs enter the brain via the LNAA transporter of the BBB, brainclearance of these amino acids is subject to the effects of theaforementioned competition. Since Phe has a relatively high affinity forthe LNAA transporter (Table 1), and since plasma levels of Phe aremarkedly elevated in phenylketonuria, PKU results in saturation of theBBB carrier protein binding sites by Phe and, hence, excessive levels ofPhe in the brain and depressed levels of the other LNAAs in the brain(Pardridge, “Blood-Brain Barrier Carrier-Mediated Transport and BrainMetabolism of Amino Acids,” Neurochem. Res., 23:635-644 (1998), which ishereby incorporated by reference).

It was thus hypothesized that non-Phe LNAA supplementation might competeeffectively with Phe at the BBB transport system, reducing Phe transportinto the brain and increasing transport of the other LNAAs and thusmoderating the symptoms of PKU (Andersen et al., “Lowering Brain PheLevels by Giving Other LNAAs,” Arch. Neurol., 33(10):684-686 (1976) andKaufman, “Phenylketonuria: Biochemical Mechanisms,” pp. 1-132 inAgranoff et al., eds, Advances in Neurochemistry, New York: Plenum Press(1977), which are hereby incorporated by reference). To reduce influx ofPhe into the brain, a supplement of branched chain neutral amino acidscomprising valine, isoleucine, and leucine, was administered to olderPKU patients (Berry, which is hereby incorporated by reference), whoreported significant improvement in behavioral deficits. Kaufmanproposed that the addition of the neurotransmitter precursors, tyrosineand tryptophan, to Berry's supplement, should lead to furtherimprovement (Kaufman, which is hereby incorporated by reference).

This hypothesis was tested experimentally by quantitative NMRmeasurement of brain levels of Phe in PKU patients during Phe oralchallenge (0.1 g/kg) with and without supplementation by 0.15 g/kgnon-Phe LNAAs (Pietz et al, “Large Neutral Amino Acids BlockPhenylalanine Transport into Brain Tissue in Patients withPhenylketonuria,” J. Clin. Invest., 103(8):1169-1178 (1999) (“Pietz”),which is hereby incorporated by reference). The LNAA supplementcontained valine, methionine, isoleucine, leucine, tyrosine, histidine,and tryptophan. Baseline plasma level of Phe was 1 mM and brain level ofPhe was 0.25 mM. Without LNAA supplementation, Pietz, which is herebyincorporated by reference, observed brain Phe increasing to 0.4 mM afterPhe challenge, accompanied by disturbed brain activity on an EEG.However, with concurrent LNAA supplementation, Phe influx into the brainwas completely blocked, and there was no slowing of EEG activity. Theseresearch studies led Nilab to develop Prekunil, a commercial LNAAsupplement for treatment of PKU.

Example 2 LNAA Supplement Formulation

As indicated above, the present inventor hypothesized that a LNAAdietary supplement designed to both compete with and suppress transportof Phe from the GI tract into the blood and to compete with and suppresstransport of Phe from the blood across the BBB into the brain could beused as a PKU treatment. More particularly, it was hypothesized thatoral administration of the LNAA supplement at each meal should suppressPhe transport from the GI tract into the blood so that the BBBtransporter system is not overwhelmed by the high levels of Phetypically present in the blood of the PKU patient.

As shown in Equation 1, the term [(aa)/Km] of each amino acid representsthat amino acid's ability to compete with Phe at a carrier proteinbinding site. As seen in Table 1, Leu, Tyr, Trp, and Met are LNAAs whichshould compete effectively with Phe at the BBB carrier protein.

Although little work has been done in characterizing the affinity of theLNAAs for the binding site of the carrier protein in the GI tract, invitro measurement of LNAA inhibition of Phe transport in humanintestinal epithelial cells (Hidalgo et al., “Transport of a LargeNeutral Amino Acid (Phenylalanine) in a Human Intestinal Epithelial CellLine: Caco-2.,” Biochim. Biophys. Acta, 1028:25-30 (1990) (“Hidalgo”),which is hereby incorporated by reference) indicated that Leu was astrong inhibitor and, interestingly, that LNAAs and basic amino acidsappear to share a carrier protein binding site in the intestinal cells,with Lys exhibiting a strong inhibition of Phe transport. Table 4 setsforth the results of experiments to determine the amino acid inhibitionof Phe transport in Caco-2 cells in which 10 μM Phe in buffer wasapplied to monolayers in presence of 1 mM concentration of each aminoacid and Phe transport across the monolayer was ratioed to that in theabsence of the competing amino acid.

TABLE 4 Inhibitor % inhibition LNAAs Leu 55% Tyr 45% Trp 36% Basic aa'sLys 50% His 33%Note that the Km value for Phe at the intestinal cell transport systemwas measured by Hidalgo, which is hereby incorporated by reference, tobe 0.56 mM, close to the value of 1 mM reported in Pardridge, which ishereby incorporated by reference. Note also that the variation in Kmbetween different LNAAs in intestinal epithelia (Table 3) is greaterthan in BBB. For example, the ratio of Km values for Phe/Leu/Met inintestinal epithelia is 1/2/5, whereas, in BBB, it is 1/1.25/1.58.

The LNAA supplement currently used for PKU treatment is Prekunil, whosecomposition, based on the amino acid makeup of human milk, is shown inTable 5. Based on the observations of inhibition of Phe transport inCaco-2 cells (Hidalgo, which is hereby incorporated by reference), theinventor of the subject invention designed alternative supplements (alsoset forth in Table 5 as SuppM1 and SuppM2) in which the Prekunil levelsof Leu and Lys were increased significantly. The increase in Leu isbelieved to further suppress Phe transport from the GI tract into theblood, and from the blood into the brain. The increase in Lys isbelieved to further suppress Phe transport from the GI tract into theblood.

TABLE 5 Prekunil SuppM1 SuppM2 amino acid (mg) (mmol) (mg) (mmol) (mg)(mmol) L-Tyr 194.1 1.07 194.1 1.07 195 1.08 L-Trp 61.1 0.30 61.1 0.30 510.25 L-Met 49.7 0.33 49.7 0.33 32 0.21 L-iLeu 31.5 0.24 31.5 0.24 350.22 L-Threo 32.8 0.28 32.8 0.28 32 0.27 L-Val 32 0.27 32 0.27 35 0.30L-Leu 30 0.24 130 1.00 80 0.61 L-His 31.3 0.20 31.3 0.20 20 0.13 L-Arg34 0.20 34 0.20 0 0 L-Lys 0 0 0 0 20 0.14 Total amino acid 496.5 3.13596.5 3.89 500 3.21 FOM-Km (app) 18.3 23.4 19.95 for Phe

Since the [Σ(aa)/Km of each aa] term of Equation 1 expresses the degreeto which each amino acid in a supplement competes with Phe at a giventransporter system, and since Km of Phe has been measured at the BBBtransport system, a figure of merit for the apparent Km for Phe in thepresence of each of the amino acid supplements of Table 5 can beexpressed by summing the ratios of the number of mmoles of each aminoacid in the supplement divided by its Km. This “figure of merit” (alsoreferred to herein as “FOM”) is a first order approximation to thedegree to which the supplement can suppress transfer of Phe from plasmainto the brain and indicates that the supplements of the subjectinvention should be 28% (SuppM1) and 9% (SuppM2) more effective insuppressing Phe transport from the plasma into the brain than Prekunil.However, note that the SuppM2 supplement is designed to optimizecompetition with Phe at the GI tract transporter, with only a smallimprovement in FOM-Km(app) for competition with Phe at the BBBtransporter.

Km values of non-Phe LNAAs at the GI tract transporter system are notknown for all the non-Phe LNAAs (Table 3), and, thus, similarcompetition terms cannot be calculated for the subject invention'ssupplements. However, under the current hypothesis, the Caco-2 cell dataof Hidalgo, which is hereby incorporated by reference, set forth inTable 4, does suggest that Leu and Lys should be effective insuppressing Phe transport out of the GI tract into the blood. Thus,augmentation of a supplement such as Prekunil with additional leucineand lysine, as in the SuppM2 supplement should increase competition withPhe at the GI tract transporter, reducing plasma Phe supply to the BBB.

Example 3 Effect of Prekunil, SuppM1, and SuppM2 on Mouse Plasma PheLevels

The supplements SuppM1 and SuppM2 were administered to mice with PKU,genotype ENU 2/2 with features of classical PKU, in single oral doses of0.5 g/kg, and the plasma phenylalanine levels were monitored at 0, 3, 6and 24 hours post-dose. It should be noted that 0.5 g/kg is a relativelylow dose of supplement as Prekunil is typically administered at 1 g/kg.It is known that LNAA supplements typically suppress phenylalanineplasma levels for several hours after ingestion, with the effect thendiminishing, such that dosing at each meal may be required. Thus, the 6hour value of phenylalanine was taken as an indicator of the degree towhich phenylalanine accumulation had been suppressed.

Data for a single mouse (P448) not receiving any supplement, a singlemouse (P455) dosed with the commercial supplement Prekunil, for a singlemouse (P430) dosed with Prekunil boosted with 35 mg Leu, for two mice(P456 and P259) dosed with Prekunil plus 100 mg Leu (i.e., SuppM1), andon two mice (P433 and P482) dosed with the SuppM2 supplement are shownin Table 6, below. More particularly, Table 6 shows the Plasma Phelevels (mg/dl) in mice administered LNAA supplements (0.5 g/kg, singledose) at 0, 3, 6, and 24 hours post-dose.

TABLE 6 Con- Prekunil + Prekunil + 100 mg Time trol Prekunil 35 mg LeuLeu (SuppM1) SuppM1 (hours) P448 P455 P430 P456 P259 P433 P482 0 33.2027.14 27.23 25.37 27.22 18.69 21.10 3 30.91 24.23 26.02 23.20 25.5714.63 16.86 6 28.91 22.17 25.21 17.49 19.02 13.68 15.39 24 30.14 20.8927.13 20.88 21.42 20.06 23.78

Boosting the Prekunil supplement with 100 mg Leu (SuppM1), an amino acidwhich should compete effectively with Phe at the binding sites ofcarrier proteins of both the intestinal epithelia and the BBB, thuseffected a significant suppression of Phe plasma level. Whereas Prekunilitself effected a 20% reduction in Phe, the SuppM1 supplement effected a30% reduction in plasma Phe at 6 hours post-dose, for both mice tested.Boosting the supplement with both leucine and lysine to target the GItransporter, as in SuppM2, effected a 70% reduction in plasma Phe at 6hours post-dose, for both mice tested. The excellent Phe suppression ofthe SuppM2 supplement is indicative of its capacity to compete moreeffectively with Phe transport at the GI transporter than the othersupplements, while maintaining the ability to compete similarly at theBBB transporter.

Plasma tyrosine analysis for the mice administered SuppM2 indicatedstable levels of 0.58-0.50 mg/dl and 0.44-0.40 mg/dl tyrosine over the24 hour experiment, for mice P433 and P482 respectively.

It will be appreciated that SuppM1 and SuppM2 may not represent optimalLNAA supplement compositions for the treatment of PKU. For example, theLNAA supplements can be modified so as to maintain required brain levelsof neurotransmitter precursor amino acids such as tyrosine andtryptophan, while improving competition of the supplement with Phe atthe GI tract transporter, and maintaining competition of the supplementwith Phe at the BBB transporter.

Example 4 Additional LNAA Supplement Formulation

An additional supplement formulation, dubbed SuppM3 is set forth inTable 7. This supplement formulation further illustrates thecompositions and methods of the present invention.

TABLE 7 amino acid SuppM3 (mg) L-Tyr 195.0 L-Trp 51.0 L-Met 32.0 L-iLeu35.0 L-Threo 32.0 L-Val 35.0 L-Leu 130.0 L-His 30.0 L-Arg 30.0 L-Lys30.0 Total amino acid 600.0

Although preferred embodiments have been depicted and described indetail herein, it will be apparent to those skilled in the relevant artthat various modifications, additions, substitutions and the like can bemade without departing from the spirit of the invention, and these aretherefore considered to be within the scope of the invention as definedin the claims which follow.

1. An LNAA supplement comprising Leu and Val or Leu and Ile, where theweight ratio of Leu to Val or Leu to Ile is greater than 2:1. The LNAAsupplement of claim 1, wherein the weight ratio of Leu to iLeu isgreater than 3:1.
 2. The LNAA supplement of claim 1, further comprisingLys.
 3. The LNAA supplement of claim 1, wherein the LNAA supplementcomprises, per 500 mg of LNAA supplement: from about 100 mg to about 290mg of Tyr; from about 25 mg to about 75 mg of Trp; from about 15 mg toabout 50 mg of Met; from about 15 mg to about 55 mg of iLeu; from about15 mg to about 50 mg of Threo; from about 15 mg to about 55 mg of Val;from about 15 mg to about 200 mg of Leu; from about 10 mg to about 30 mgof His; and from about 5 mg to about 200 mg of Lys.
 4. The LNAAsupplement of claim 1, wherein the LNAA supplement comprises, per 500 mgof LNAA supplement, from about 10 mg to about 30 mg of Lys.
 5. The LNAAsupplement of claim 1, comprising, per 600 mg of LNAA supplement: fromabout 100 mg to about 290 mg of Tyr; from about 30 mg to about 90 mg ofTrp; from about 25 mg to about 75 mg of Met; from about 15 mg to about45 mg of iLeu; from about 15 mg to about 50 mg of Threo; from about 15mg to about 50 mg of Val; from about 40 mg to about 200 mg of Leu; fromabout 15 mg to about 45 mg of His; and from about 15 mg to about 50 mgof Arg.
 6. The LNAA supplement of claim 1, wherein the LNAA supplementfurther comprises, per 600 mg of LNAA supplement, from about 5 mg toabout 200 mg of Lys.
 7. A method for treating a subject suffering from adeficiency of an LNAA, comprising enterally administering to the subjectthe LNAA supplement of claim
 1. 8. The method of claim 7, where thecondition is phenylketonuria and/or phenylalanemia.
 9. The method ofclaim 7, where the condition is tyrosinemia.
 10. The method of claim 7,where the condition is alkaptonuria.
 11. The method of claim 7, wheresaid enteral administration is carried out substantially at mealtime.12. The method of claim 7, where said enteral administration is oraladministration.